The use of microlevers (or standard AFM cantilevers, combined with detection by an external optical lever involving a laser or a photodiode, for chemical force spectroscopy in the analysis of molecular bonds has already been demonstrated and systematically applied (Janshoff et al., 2000, Angew. Chem. Int. Ed., Vol 39, pages 3212-3237).
Several patents and patent applications relate to chemical force spectroscopy using an AFM tip. U.S. Pat. No. 7,013,717 in the name of Veeco Instruments Inc., contemplates the grafting of single-stranded DNA on the tip of an AFM cantilever and of a complementary single-stranded DNA on the probed surface. International Application WO 2007/087653 in the name of the University of Linz, proposes a method for detecting 5-methylcytosine involving anti-5-methylcytosine antibodies grafted on the tip of an AFM cantilever.
Further, certain recent commercial chemical force spectroscopy systems begin to appear on the market like the automated machine proposed by the nAmbition or JPK companies.
In parallel, the use of AFM probes with an integrated piezo-electric sensor has already been demonstrated by several teams worldwide.
In particular, in U.S. Pat. No. 5,641,896, Karraï uses, for applications in near-field optical microscopy (or SNOM for Scanning Near-field Optical Microscopy), the force return provided by the piezo-electric probe for approaching an optical fiber adhesively bonded on the probe at a few tens of nanometers from the surface. U.S. Pat. No. 6,240,771 describes the use of such an AFM probe with an integrated piezo-electric sensor with a metal, well sharpened tip, adhered to the end of the probe in order to scan crystals with atomic resolution. Kaghesima et al. (2002, Applied Surface Science, Vol. 188, pages 440-444) and Rensen and van Hulst (2000, Appl. Phys. Lett., Vol. 77, pages 1557-1559) have also demonstrated the use of this probe for AFM in a liquid medium with total immersion of the probe, in an organic solution or in ultrapure water respectively. Other teams have used the piezo-electric probe outside the liquid, but with the tip immersed in a liquid meniscus for probing the surface below (for example Koopman et al., 2003, Appl. Phys. Lett., Vol. 83, pages 5083-5085).
Finally, some studies have illustrated the use of these probes with integrated piezo-electric sensor for analyzing the properties of liquids such as viscosity and density (U.S. Pat. No. 6,393,895 in the name of Symyx Technologies Inc.) or as a chemical nose allowing detection of selectively grafted molecules (U.S. Pat. No. 6,393,895 and patent application US 2010/0068697 in the name of Drexel University).
Probes with an integrated sensor like piezo-electric sensors have several advantages as compared with a standard AFM cantilever since they give the possibility of getting rid of the conventional optical detection system of the latter. They are easier to integrate and to use in extreme environments. But an essential point as to their use in biological applications is the requirement of their being able to be immersed in a biological solution while retaining good mechanical and electrical behavior. This condition is not obvious to meet because the saline solutions used in biology such as HEPES, Tris or PBS are extremely conductive because of the presence of many ions in the aqueous medium. Thus, the probe cannot be used as such in these liquids because of inevitable problems of electric leakages, short-circuits or corrosion effects on the electrodes of the piezo-electric sensor.
The inventors therefore set themselves the goal of proposing a system comprising an AFM probe and notably an AFM probe with an integrated piezo-electric sensor useful for analyzing molecular bonds in chemical force spectroscopy and this, in buffer solutions, notably in solutions applied in the field of biology. The AFM probe should further retain good mechanical and electrical properties, have a good quality factor, when the latter is used in buffer solutions and be obtained by a method simple to apply. As a reminder, the quality factor is a description of the damping of an oscillating system. It is given by the ratio between the transmitted energy and the lost energy. In the case of a probe, the quality factor represents the energy transmitted in the pass band around the resonance relatively to the energy contained outside the pass band.